Original Article
Effects of Muntingia calabura L. on
isoproterenol-induced myocardial
infarction
Nivethetha M, Jayasri J, Brindha P
ABSTRACT
Introduction : This study was designed to
scientifically evaluate the effects of an aqueous
extract of Muntingia calabura L. (M. calabura),
a medicinal herb, on isoproterenol-induced
myocardial infarction (MI) in rat models.
Methods: Six groups of Wistar albino rats, each
comprising six animals, were selected for this study.
Group I served as a control, Group II rats were given
isoproterenol (20 mg/100g, subcutaneously), and
Group III rats were given M. calabura leaf extract
(300 mg/kg). Groups IV, V and VI rats were given
M. calabura leaf extract (100 mg/kg, 200 mg/kg and
300 mg/kg, respectively) and isoproterenol (20
mg/100g subcutaneously) prior to MI induction.
The transaminases (aspartate transaminase and
alanine transaminase), lactate dehydrogenase
(LDH) and creatine phosphokinase (CK), were
estimated in both the serum and heart tissues, and
the serum uric acid level was also estimated.
Results: Isoproterenol significantly increased
the activities of CK, LDH and the transaminases
in serum with a concomitant decrease in these
enzymes in tissue. Pretreatment with the aqueous
leaf extract of M. calabura at a dose of 300 mg/kg
body weight for 30 days had a significant effect
on the activities of marker enzymes compared
to the other groups. Serum uric acid level, which
Department of increased on isoproterenol administration,
Biochemistry,
Srimad Andavan Arts registered near normal values on treatment with
and Science College,
the leaf extract under study.
Tiruchirappalli
620005,
Tamil Nadu,
India Conclusion: The study confirms the protective
effects of M . calabura leaf extract against
Nivethetha M, MSc,
MPhil isoproterenol-induced biochemical alterations
Lecturer
in rats.
Jayasri J, MSc
Research Scholar
Keywords: isoproterenol, leaf extract, Muntingia
Brindha P, MSc, PhD
Dean calabura, myocardial infarction, uric acid
Correspondence to: Singapore Med J 2009; 50(3): 300-302
Dr Pemiah Brindha
Tel: (91) 431 223 1938
Fax: (91) 431 223 1938 Introduction
Email: brindhajana@
yahoo.com The entire world population is turning towards natural
Singapore Med J 2009; 50 (3) : 300
drugs because of the widespread belief that “green
medicines” are healthier and safer than synthetic ones.(1)
It is also gaining greater acceptance from the public
and the medical profession due to greater advances in
understanding the mechanism of action by which herbs can
positively influence health and quality of life.(2) Globally,
myocardial infarction (MI) is one of the leading causes of
death for both men and women.(3) Due to changing lifestyles
in developing countries, such as India, and particularly
in urban areas, MI is making an increasingly important
contribution to mortality statistics.(4) MI is a complex
phenomenon affecting the mechanical, electrical, structural
and biochemical properties of the heart.(5) Although
modern drugs are effective in preventing cardiovascular
disorders, their use is often limited because of their side
effects.(6) Many plant sources such as Commiphora mukul,
Terminalia arjuna, Inula racemosa, Coleus forskohlii
and their extracts are being used extensively to treat heart
ailments. Muntingia calabura L. (M. calabura), which
belongs to the family Elaeocarpaceae, is popularly known
for its antiseptic and antispasmodic properties besides being
a proven hypotensive drug.(7)
Several methods have been used to study the
beneficial effects of many drugs on cardiac function.(8) The
administration of isoproterenol, a β-adrenergic agonist,
has been found to cause severe stress in the myocardium,
resulting in the infarct-like necrosis of the heart muscle.(9)
Isoproterenol-induced MI serves as a well-standardised
model because the pathophysiological changes following
isoproterenol administration are comparable to those taking
place in human MI.(10) The present communication thus
embodies the cardioprotective effect of the M. calabura leaf
extract on myocardial necrosis induced by isoproterenol
with reference to marker enzymes in the serum, heart tissues
and serum uric acid.
Methods
Adult male albino rats of the Wistar strain weighing 120–
150 g were selected for the study. The rats were fed with
commercial pelleted rat chow bought from Sai Durga Feeds
and Foods, Bangalore, India and water ad libitum. The
animals were housed in clean polypropylene cages lined
with husk, changed every 24 hours under a 12-hour light/
dark cycle at 25°C. The study was carried out after obtaining
 Singapore Med J 2009; 50 (3) : 301

Table I. Effect of M. calabura pretreatment on isoproterenol-induced changes in the activities of serum AST, ALT,
CK, LDH and uric acid.
Group Aspartate transaminase¶ Alanine transaminase¶ Creatine kinase** Lactate dehydrogenase¶ Uric acid

I 31.77 ± 0.32 14.62 ± 0.16 274.75 ± 0.21 78.8 ± 0.21 3.87 ± 0.09
II 52.63 ± 0.38* 25.68 ± 0.23* 530.52 ± 0.31* 145.43 ± 0.17* 5.55 ± 0.15*
III 32.33 ± 0.33 13.75 ± 0.16 272.43 ± 0.28 78.23 ± 0.10 4.03 ± 0.07
IV 36.77 ± 0.18† 15.67 ± 0.2† 283.45 ± 0.28† 81.0 ± 0.22† 4.5 ± 0.13†
V 35.37 ± 0.18‡ 15.42 ± 0.17‡ 279.70 ± 0.22‡ 80.95 ± 0.15‡ 4.33 ± 0.20‡
VI 33.65 ± 0.15§ 14.27 ± 0.19§ 275.57 ± 0.18§ 80.23 ± 0.43§ 3.93 ± 0.13§

Data is expressed as mean ± SEM for the six animals in each group.
*p < 0.001 compared with Group I. †,‡,§ p < 0.001 compared with Group II.
¶
nmol of pyruvate liberated/sec/mg protein.
** µmol of phosphorous liberated/sec/mg protein.

Table II. Effect of M. calabura pretreatment on isoproterenol-induced changes in the activities of membrane AST,
ALT, CK and LDH.
Group Aspartate transaminase¶ Alanine transaminase¶ Creatine kinase** Lactate dehydrogenase¶

I 43.51 ± 0.20 25.28 ± 0.15 13.13 ± 0.10 117.22 ± 0.15

II 27.9 ± 0.13* 16.45 ± 0.16* 9.45 ± 0.13* 80.43 ± 0.18*
III 44.46 ± 0.14 26.18 ± 0.16 14.55 ± 0.13 115.43 ± 0.20
IV 42.18 ± 0.30† 25.35 ± 0.15† 12.36 ± 0.12† 114.78 ± 0.21†
V 44.48 ± 0.15‡ 25.92 ± 0.17‡ 13.48 ± 0.12‡ 115.38 ± 0.14‡
VI 45.45 ± 0.23§ 27.25 ± 0.20§ 14.16 ± 0.07§ 116.85 ± 0.26§

Data is expressed as mean ± SEM for the six animals in each group.
*p < 0.001 compared with Group I. †,‡,§ p < 0.001 compared with Group II.
¶
nmol of pyruvate liberated/sec/mg protein.
** µmol of phosphorous liberated/sec/mg protein.

the necessary approval from the Institutional Animal Ethics
Committee. Isoproterenol and adenosine triphosphate
were obtained from Sigma Chemical Company, St. Louis,
MO, USA, and all other chemicals used were of analytical
grade. The plant M. calabura L. was identified using the
flora of Gamble and authenticated with a voucher specimen
deposited in the RAPINET Herbarium, St Joseph’s College,
Tiruchirappalli. An aqueous extract was prepared and
suspended in 0.9% saline and used for the study.
The rats were divided into six groups of six animals
each. Group I served as a control, Group II rats were
administered with isoproterenol (20 mg/100g administered
subcutaneously twice at an interval of 24 h) dissolved in
normal saline.(6) Group III rats were pretreated with M.
calabura leaf extract (300 mg/kg) for a period of 30 days.
Groups IV, V and VI animals were pretreated with M.
calabura leaf extract (100 mg/kg, 200 mg/kg and 300 mg/
kg, respectively) for a period of 30 days(11)
and isoproterenol
(20 mg/100g subcutaneously twice at an interval of 24
hours) at the end of the treatment period on the 29th and
30th days.
After the experimental period, the rats were sacrificed
by cervical decapitation. Blood was collected and the serum
was separated and used for the assay of marker enzymes
lactate dehydrogenase (LDH) by the method of King,(12)
aspartate transaminase (AST) and alanine transaminase
(ALT) were estimated according to the method of Mohun
and Cooke(13) and creatine kinase (CK) by the method of
Okinaka et al.(14) The serum was also used for the assay
of marker enzymes as well as uric acid, and protein was
estimated by the methods of Caraway(15) and Lowry et al,(16)
respectively. The heart was dissected, immediately washed
in ice-cold saline and a homogenate was prepared in 0.1 M
Tris-HCl buffer (pH 7.4). The homogenate was centrifuged
and the supernatant was used for the assay of marker
enzymes. Student’s t-test was used for statistical analysis.
Values are expressed as the mean ± standard error of the
mean (SEM) for the six animals in each group. A value of p
< 0.001 was considered statistically significant.
Results
There was a significant elevation in the transaminases
(AST and ALT), CK and LDH activities in isoproterenol-
injected animals compared to the controls (Table I). In the
Groups IV, V and VI rats pretreated with the M. calabura
leaf extract, there was a significant reduction in the level
of uric acid and the activity of marker enzymes compared
with the isoproterenol-administered rats (Group II).
Rats in Group II were given isoproterenol
subcutaneously. Rats in Groups IV, V and VI were given
M. calabura leaf extract and isoproterenol subcutaneously
at the end of the treatment period. Compared to controls,
there was a significant reduction in the activity of marker
enzymes (AST, ALT, CK and LDH) on isoproterenol

administration (Group II) (Table II). Pretreatment with
M. calabura extract (Groups IV, V and VI) retained the
activity of these enzymes to near normal levels. In all the
parameters studied, M. calabura extract at a dose of 100
mg/kg showed a minor effect, whereas doses of 200 mg/kg
and 300 mg/kg showed significant effects, with the dose of
300 mg/kg found to be the most effective.
Discussion
Medicinal plants have long been valued as sources of new
compounds with cardioprotective activity. The present study
demonstrated that the M. calabura leaf extract has efficiently
protected the myocardium against isoproterenol-induced
MI. Isoproterenol administration brought about a significant
decrease in the activities of cardiac marker enzymes such as
AST, ALT, CK and LDH in the myocardial tissue (Table II),
with a subsequent increase in the activities of these enzymes
in the serum. This might be due to the damage in the heart
muscle, rendering the leakage of enzymes into the serum.(17)
The significant rise observed in the levels of diagnostic
marker enzymes in the serum of Group II isoproterenol-
administered rats as compared to that of Group I control
rats (Table I) is an indication of the severity of the necrotic
damage to the myocardial membrane.(18) Enzymes are the
best markers of tissue damage because of their specificity
and catalytic activity to the tissue.(19) The release of cellular
enzymes reflects non-specific alterations in the membrane
integrity and permeability as a response to β-adrenergic
stimulation.
In the present study, pretreatment with the M. calabura
leaf extract significantly prevented isopreterenol-induced
elevation in the levels of the diagnostic marker enzymes
of Groups IV, V and VI animals compared to Group II rats,
probably due to the protective effect of the M. calabura
leaf extract on the myocardium; this reduced the extent of
myocardial damage and thereby restricted the leakage of
these enzymes from the myocardium.(20) The significant
elevation observed in the level of serum uric acid in the
isoproterenol-injected groups could be due to the excessive
degradation of purine nucleotides and proteolysis.(21) In
conclusion, the results of the present study indicate that
the prior administration of the M. calabura leaf extract
attenuates isoproterenol-induced MI. The cardioprotective
effect of the M calabura leaf extract is probably related to
its ability to strengthen the myocardial membrane by its
membrane-stabilising action. M. calabura leaf extract at
the dose level of 300 mg/kg was found to be more effective
than the other dose levels. In this study, the cardioprotective
potential of M. calabura leaf extract is evident.
Singapore Med J 2009; 50 (3) : 302
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